Infectious causes of cholesteatoma and treatment of infected ossicles prior to reimplantation by hydrostatic high-pressure inactivation.

Chronic inflammation, which is caused by recurrent infections, is one of the factors contributing to the pathogenesis of cholesteatoma. If reimplantation of autologous ossicles after a surgical intervention is intended, inactivation of planktonic bacteria and biofilms is desirable. High hydrostatic pressure treatment is a procedure, which has been used to inactivate cholesteatoma cells on ossicles. Here we discuss the potential inactivating effect of high hydrostatic pressure on microbial pathogens including biofilms. Recent experimental data suggest an incomplete inactivation at a pressure level, which is tolerable for the bone substance of ossicles and results at least in a considerable reduction of pathogen load. Further studies are necessary to access how far this quantitative reduction of pathogens is sufficient to prevent ongoing chronic infections, for example, due to forming of biofilms.

Effects of high hydrostatic pressure on bacterial growth on human ossicles explanted from cholesteatoma patients.

BACKGROUND: High hydrostatic pressure (HHP) treatment can eliminate cholesteatoma cells from explanted human ossicles prior to re-insertion. We analyzed the effects of HHP treatment on the microbial flora on ossicles and on the planktonic and biofilm states of selected isolates. METHODOLOGY: Twenty-six ossicles were explanted from cholesteatoma patients. Five ossicles were directly analyzed for microbial growth without further treatment. Fifteen ossicles were cut into two pieces. One piece was exposed to HHP of 350 MPa for 10 minutes. Both the treated and untreated (control) pieces were then assessed semi-quantitatively. Three ossicles were cut into two pieces and exposed to identical pressure conditions with or without the addition of one of two different combinations of antibiotics to the medium. Differential effects of 10-minute in vitro exposure of planktonic and biofilm bacteria to pressures of 100 MPa, 250 MPa, 400 MPa and 540 MPa in isotonic and hypotonic media were analyzed using two patient isolates of Staphylococcus epidermidis and Neisseria subflava. Bacterial cell inactivation and biofilm destruction were assessed by colony counting and electron microscopy. PRINCIPAL FINDINGS: A variety of microorganisms were isolated from the ossicles. Irrespective of the medium, HHP treatment at 350 MPa for 10 minutes led to satisfying but incomplete inactivation especially of gram-negative bacteria. The addition of antibiotics increased the efficacy of elimination. A comparison of HHP treatment of planktonic and biofilm cells showed that the effects of HPP were reduced by about one decadic logarithmic unit when HPP was applied to biofilms. High hydrostatic pressure conditions that are suitable to inactivate cholesteatoma cells fail to completely sterilize ossicles even if antibiotics are added. As a result of the reduced microbial load and the viability loss of surviving bacteria, however, there is a lower risk of re-infection after re-insertion.

An important procedure in ossiculoplasty: autoclaving the ossicles.

Autoclaving of the ossicles prior to ossiculoplasty is a very important procedure in surgery of cholesteatomatous chronic otitis media. Autoclaving allows the reuse of the ossicles removed from patients with cholesteatomatous chronic otitis media as an autograft. It also allows utilization of the malleus, incus and stapes taken from cadavers or of the uneroded malleus and incus removed from patients undergoing non-functional middle ear surgery chosen carefully with detailed history and laboratory analysis. The powerful disinfecting effect of the sterilising procedures of the homografts inactivates prion proteins which cause degenerative encephalopathies. In various studies, it has been concluded that autoclaving does not alter the matrix of the bone which is responsiable for its biophysical properties, whereas it removes all viable cells within the bone and dentures the soft tissue attached to the surface of the ossicle. We have also found confirming histological results in our two previous studies published in 1999 and 2001. It is usually recommended that soft tissue on the surface of the ossicles is removed before autoclaving. It is also sufficient to autoclave the ossicles at 134 degrees C, and at 2.5 atmosphere pressure for 5 minutes in a flash autoclave. The autoclaving time of the homograft ossicles must be longer, for 20 minutes. Following the autoclaving, these homograft ossicles should be kept at pH 5.6 for 3 days, then use a solution of 0.5% formaldehyde at pH 7 and 4 degrees C for 21 days and it should be washed with physiologic saline solution three times for 7 minutes before use. Depending on the results obtained from our planned comparative experimental study, there will be no need for additional formaldehyde fixation procedure after autoclaving and the autoclaved ossicles will be used immediately without preservation in formaldehyde solution for 24 days.

[Are cialit-preserved ossicles as middle ear implants still bacteriologically defensible? Their replacement by pre-formed implants fashioned from teeth]. / Sind Cialit-konservierte Ossikel als Mittelohrimplantate bakteriologisch noch vertretbar? Ihr Ersatz durch vorgefertigte Implantate aus Zähnen.

Recent studies have demonstrated a limited bactericidal activity of Cialit in the preservation of otologic transplants. The authors therefore performed several tests following routine treatment of the grafts. First, the contamination of allogenic ossicles was studied; afterwards, the bactericidal activity of Cialit was tested against standardized germs and clinical isolates. At the highly toxic concentration of 1:2000, Cialit is bactericidal. For clinical purposes, however, the less toxic solution of 1:5000 is generally used, which is not sufficiently safe against certain resistant clinical isolates. The fact that the activity of Cialit depends on its concentration explains the controversial findings in the literature. Furthermore, it is doubtful whether a weaker solution would prevent transmission of an HIV or hepatitis infection. Therefore, the authors recommend the autoclaving of otologic allografts. This also has forensic reasons; for example if the serologic status of the donor is unknown. As ossicles become brittle under this procedure, the authors now use sculptured parts of teeth, in which the hydroxyl-apatite is much more compact. The preformed prostheses can be autoclaved without problem. More than 150 reconstructions have proved the acceptance of this cheap implant material. Furthermore, forensic problems with ossicles from corpses do not arise.

[Transmissibility of HIV infections via allogeneic transplants]. / Zur Ubertragbarketi von HIV-Infektionen durch allogene Transplantate.

HIV-infections can be transferred by blood and blood products, semen or organ transplants. For this reason, the WHO recommended a screening of all donors of blood, semen, tissues and organs. Those who belong to high-risk groups should be excluded from donations. If this recommendation should be also applied to cialit preserved auditory aussicles, this would probably mean the end of allogeneic transplantations of middle-ear surgery which are being performed daily. Therefore, we carried out experimental examinations with HIV-infected auditory aussicles in cialit. According to our results, it can be assumed that a virus-destruction takes place via a cialit-treatment. The same applies to solvent-dried fascia as to auditory aussicles which have been preserved in the same way. Consequently, there is no objection to further application of these allogeneic transplants in respect to HIV-infections.

[Bacteriologic studies on the preservation of bone implants]. / Bakteriologische Untersuchungen zur Konservierung von Knochenimplantaten.

High demands on freedom from germs are made on auditory ossicles to be used as implants. Postoperative infections often result from the existence of germ-contaminated ossicles. That is why we undertook microbiological examinations, using three frequently applied preserving methods for auditory ossicles kept for tympanoplasties. The results of the examinations were: Disinfection by Cialit (often used in clinical practice) produced unsatisfactory results. Disinfection by ethyl alcohol (70 per cent) and a subsequent treatment with thiomersal-Ringer's solution (0.4 per cent) yielded satisfactory results. Disinfection by preservation with an aqueous solution of formalin and subsequent treatment with Cialit proved most effective. None of the applicated germs could be regrown.